Generation of Rab-Based Transgenic Lines for In Vivo Studies of Endosome Biology in Zebrafish

被引：77

作者：

Clark, Brian S. ^{[2
]}

Winter, Mark ^{[1
]}

Cohen, Andrew R. ^{[1
]}

Link, Brian A. ^{[2
]}

机构：

[1] Univ Wisconsin, Dept Elect Engn & Comp Sci, Milwaukee, WI 53211 USA

[2] Med Coll Wisconsin, Dept Cell Biol Neurobiol & Anat, Milwaukee, WI 53226 USA

来源：

DEVELOPMENTAL DYNAMICS | 2011年 / 240卷 / 11期

基金：

美国国家卫生研究院;

关键词：

endosome dynamics; neuroepithelia; organelle polarization; zebrafish transgenic lines; GDP DISSOCIATION INHIBITOR; TARGETED GENE-EXPRESSION; FGF8 MORPHOGEN GRADIENT; GTP-BINDING-PROTEINS; ENDOCYTIC PATHWAY; MOLECULAR-CLONING; HOMEOBOX GENE; MEMBRANE; DISEASE; CELLS;

D O I：

10.1002/dvdy.22758

中图分类号：

R602 [外科病理学、解剖学]; R32 [人体形态学];

学科分类号：

100101 ;

摘要：

The Rab family of small GTPases function as molecular switches regulating membrane and protein trafficking. Individual Rab isoforms define and are required for specific endosomal compartments. To facilitate in vivo investigation of specific Rab proteins, and endosome biology in general, we have generated transgenic zebrafish lines to mark and manipulate Rab proteins. We also developed software to track and quantify endosome dynamics within time-lapse movies. The established transgenic lines ubiquitously express EGFP fusions of Rab5c (early endosomes), Rab11a (recycling endosomes), and Rab7 (late endosomes) to study localization and dynamics during development. Additionally, we generated UAS-based transgenic lines expressing constitutive active (CA) and dominant-negative (DN) versions for each of these Rab proteins. Predicted localization and functional consequences for each line were verified through a variety of assays, including lipophilic dye uptake and Crumbs2a localization. In summary, we have established a toolset for in vivo analyses of endosome dynamics and functions. Developmental Dynamics 240: 2452-2465, 2011. (C) 2011 Wiley Periodicals, Inc.

引用

页码：2452 / 2465

页数：14

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